In situ formation of H2O2 for P450 peroxygenases.

In situ formation of H2O2 for P450 peroxygenases.

Bioorganic & medicinal chemistry (2014-07-06)

Caroline E Paul, Ekaterina Churakova, Elmer Maurits, Marco Girhard, Vlada B Urlacher, Frank Hollmann

ABSTRACT

An in situ H2O2 generation approach to promote P450 peroxygenases catalysis was developed through the use of the nicotinamide cofactor analogue 1-benzyl-1,4-dihydronicotinamide (BNAH) and flavin mononucleotide (FMN). Final productivity could be enhanced due to higher enzyme stability at low H2O2 concentrations. The H2O2 generation represented the rate-limiting step, however it could be easily controlled by varying both FMN and BNAH concentrations. Further characterization can result in an optimized ratio of FMN/BNAH/O2/biocatalyst enabling high reaction rates while minimizing H2O2-related inactivation of the enzyme.

MATERIALS

Product Number

Brand

Product Description

70082

Sigma-Aldrich

Myristic acid, ≥98.0% (GC)

93440

Supelco

Trizma^® base, reference material for titrimetry, certified by BAM, >99.5%

92360

Sigma-Aldrich

Chlorotrimethylsilane, puriss., ≥99.0% (GC)

92361

Sigma-Aldrich

Chlorotrimethylsilane, ≥98.0% (GC)

93352

Sigma-Aldrich

Trizma^® base, ≥99.0% (T)

93362

Sigma-Aldrich

Trizma^® base, BioUltra, for molecular biology, ≥99.8% (T)

386529

Sigma-Aldrich

Chlorotrimethylsilane, purified by redistillation, ≥99%

252859

Sigma-Aldrich

Tris(hydroxymethyl)aminomethane, ACS reagent, ≥99.8%

179272

Sigma-Aldrich

Diethyl ether

T4661

Sigma-Aldrich

Trizma^® base, ≥99.9% (titration), crystalline

T6791

Sigma-Aldrich

Trizma^® base, BioXtra, pH 10.5-12.0 (1 M in H₂O), ≥99.9% (titration)

T6066

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Trizma^® base, BioPerformance Certified, meets EP, USP testing specifications, suitable for cell culture, ≥99.9% (titration)

T6687

Sigma-Aldrich

Tromethamine, meets USP testing specifications

T1503

Sigma-Aldrich

Trizma^® base, Primary Standard and Buffer, ≥99.9% (titration), crystalline

93350

Sigma-Aldrich

Trizma^® base, puriss. p.a., ≥99.7% (T)

95541

Sigma-Aldrich

Chlorotrimethylsilane, produced by Wacker Chemie AG, Burghausen, Germany, ≥99.0% (GC)

89595

Supelco

Chlorotrimethylsilane, for GC derivatization, LiChropur^™, ≥99.0% (GC)

PHR1347

Supelco

Tromethamine, pharmaceutical secondary standard, certified reference material

T2550000

Trometamol, European Pharmacopoeia (EP) Reference Standard

RDD008

Sigma-Aldrich

Trizma^® base, anhydrous, free-flowing, Redi-Dri^™, ≥99.9%

676845

Sigma-Aldrich

Diethyl ether, ACS reagent, ≥98.0%, contains ≤2% ethanol and ≤10ppm BHT as inhibitor

24004

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Diethyl ether, puriss., contains ~5 mg/L 2,6-di-tert.-butyl-4-methylphenol as stabilizer, meets analytical specification of Ph. Eur., BP, ≥99.5% (GC)

V900483

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Trizma^® base, Vetec^™, reagent grade, ≥99%

58958

Supelco

Ethyl acetate, analytical standard

41639

Sigma-Aldrich

Dimethyl sulfoxide, BioUltra, for molecular biology, ≥99.5% (GC)

D2438

Sigma-Aldrich

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